The actin-bundling protein TRIOBP-4 and -5 promotes the motility of pancreatic cancer cells
Introduction
Cell migration is a fundamental cellular process that is important for many physiological functions including embryogenesis, immune response, and tissue repair. It is a coordinated procedure depending on the dynamic actin cytoskeleton reorganization via the formation of specialized cellular structures controlled by actin-binding proteins [1], [2]. Whereas cell migration is important for normal cellular functions, it can be deregulated in cancer cells and contribute to their invasive behaviors. In fact, aberrant expression and biochemical activities of actin-bundling proteins have been reported to correlate with the invasiveness and chemotherapy-resistance of many tumors [3], [4].
At the leading edge of motile cells, actin filaments are assembled into tight parallel actin bundles by actin-bundling proteins to form the finger-like protrusive structures called filopodia. This actin-enriched specialized structure functions as a sensor to probe the extracellular environment and plays a key role in promoting cell motility [5], [6]. The abundance of filopodia has been demonstrated as an indicator for cancer cell invasiveness [7]. Fascin is the major actin-bundling protein identified in filopodia [8], [9] and its role in various cancers has been studied. Up-regulation of fascin has been reported in cancers of liver, ovary, prostate, colon, and pancreas, and is correlated with cancer invasiveness [7], [10], [11], [12], [13], [14]. In these cancers, fascin is shown to promote filopodial formation and motility of the tumor cells.
The guanine nucleotide exchange factor (GEF) trio binding protein isoforms 4 and 5 (TRIOBP-4/-5) is an F-actin-bundling protein originally found to be associated with hearing loss both in humans and mice [15], [16], [17]. The human and mouse TRIOBP protein is largely classified into three isoforms with a long isoform (TRIOBP-5) and two shorter isoforms equivalent to the N-terminus and C-terminus of TRIOBP-5, which are named TRIOBP-4 and TRIOBP-1 (or Tara), respectively. There are no common sequences between TRIOBP-1 and -4. TRIOBP-1 is ubiquitously expressed and has been shown to regulate adherens junctions and actin cytoskeleton reorganization mostly in stress fibers and cortical F-actin [18], [19]. In contrast, TRIOBP-4 and -5 are predominantly expressed in the inner ear and the retina of normal adult tissues, where they are believed to provide appropriate durability and rigidity for stereocilia of hair cells for normal hearing [17]. Interestingly, different from fascin, which is intercalated between actin filaments via its two major actin-binding sites [20], [21], [22], TRIOBP-4 and -5 form extremely dense F-actin bundles without detectable inter-filament spaces, raising the possibility that they more likely wrap around actin bundles [17], [23]. To date, the biological functions of TRIOBP-4/-5 have not been revealed in any other tissues except in inner ear hair cells for hearing.
Here, we showed that TRIOBP-4 and -5 are significantly up-regulated in pancreatic cancer cells and human cancer tissues. Moreover, knockdown of TRIOBP-4 and -5 inhibited the filopodial formation and motility of pancreatic cancer cells. Our study reveals a novel regulator for pancreatic cancer migration, which may be responsible for the aggressive biology of pancreatic cancer cells.
Section snippets
siRNAs and quantitative real-time PCR
The siRNAs against human TRIOBP-4 and -5 (T4 + 5 siRNA) were synthesized by Integrated DNA Technologies and their targeting sequences were listed in Supplemental Table S1. Non-targeting RNA duplexes were used as negative controls (control siRNA, Integrated DNA Technologies). Total RNA from cultured cells was extracted using RNeasy Kit (QIAGEN), and the first strand cDNA was synthesized using Superscript III reverse transcriptase and random hexamers as primers (Life Technologies). Quantitative
TRIOBP-4 and -5 are up-regulated in PDAC cancer cells
Using isoform-specific primers (Fig. 1A), we examined the mRNA expression of TRIOBP isoforms in normal pancreatic ductal epithelial cells (HPDE) and pancreatic cancer cell lines by RT-PCR (Fig. 1B). Compared with HPDE, TRIOBP-4 was significantly overexpressed in HPAC cells, and TRIOBP-5 was exclusively upregulated in PANC-1 and MIA PaCa-2 cells, while Colo-357 cells showed a higher level of both TRIOBP-4 and -5. By contrast, TRIOBP-1 expression level was not significantly changed among all the
Discussion
In this study, we found that the actin-bundling protein TRIOBP-4/-5 is up-regulated in several human pancreatic cancer cells and tissues. We further demonstrated that TRIOBP-4/-5 promotes pancreatic cancer cell migration via the regulation of actin cytoskeleton reorganization in the filopodia of the cells. Therefore, we propose that TRIOBP-4/-5 may be a novel regulator for the aggressive behavior of pancreatic cancer.
PDAC originates from the ductal cells of exocrine pancreas and is the most
Conflicts of interest
To the best of our knowledge, no conflict of interest, financial or other, exists.
Acknowledgements
We acknowledge the professional supports of the Microscope, Imaging and Cytometry Resources Core (MICR) of Wayne State University School of Medicine and the Laboratory of Analytical Electron Microscopy (LAEM), Department of Chemistry, Wayne State University. We thank Dr. Avraham Raz (Karmanos Cancer Institute, Wayne State University, Detroit, MI) for the comments and suggestions. We thank Dr. Thomas B. Friedman (NIDCD, NIH, Rockville, MD) for providing the antibody against TRIOBP-4 and -5. This
References (33)
- et al.
Building distinct actin filament networks in a common cytoplasm
Curr. Biol
(2011) - et al.
The physics of filopodial protrusion
Biophys. J.
(2005) - et al.
Formation of filopodia in coelomocytes: localization of fascin, a 58,000 dalton actin cross-linking protein
Cell
(1979) - et al.
Mutations in TRIOBP, which encodes a putative cytoskeletal-organizing protein, are associated with nonsyndromic recessive deafness
Am. J. Hum. Genet
(2006) - et al.
Mutations in a novel isoform of TRIOBP that encodes a filamentous-actin binding protein are responsible for DFNB28 recessive nonsyndromic hearing loss
Am. J. Hum. Genet
(2006) - et al.
Actin-bundling protein TRIOBP forms resilient rootlets of hair cell stereocilia essential for hearing
Cell
(2010) - et al.
Mechanism of actin filament bundling by fascin
J. Biol. Chem
(2011) - et al.
Molecular mechanism of fascin function in filopodial formation
J. Biol. Chem
(2013) - et al.
Identification of an actin binding region and a protein kinase C phosphorylation site on human fascin
J. Biol. Chem
(1997) - et al.
CXCR2 macromolecular complex in pancreatic cancer: a potential therapeutic target in tumor growth
Transl. Oncol
(2013)
The actin-bundling protein fascin stabilizes actin in invadopodia and potentiates protrusive invasion
Curr. Biol
The E3 ubiquitin ligase HECTD3 regulates ubiquitination and degradation of Tara
Biochem. Biophys. Res. Commun
Actin, a central player in cell shape and movement
Science
Actin binding proteins: their ups and downs in metastatic life
Cell. Adh. Migr
Actin-bundling proteins in cancer progression at a glance
J. Cell Sci
Filopodia: molecular architecture and cellular functions
Nat. Rev. Mol. Cell Biol
Cited by (15)
Sp1 Suppresses miR-3178 to Promote the Metastasis Invasion Cascade via Upregulation of TRIOBP
2018, Molecular Therapy Nucleic AcidsCitation Excerpt :Primers used in all experiments were synthesized at Comate Biosciences (Jilin, China), and miR-3178 mimics and according inhibitors were purchased from Ribio Company (Guangzhou, China). Specific primers for TRIOBP-1, TRIOBP-4, and TRIOBP-5 were designed with reference.43 The primer and oligo sequences are listed in Table 1.
Electroactive nanoinjection platform for intracellular delivery and gene silencing
2023, Journal of NanobiotechnologySilicon-Nanotube-Mediated Intracellular Delivery Enables Ex Vivo Gene Editing
2020, Advanced Materials